Device for regeneration of articular cartilage and other tissue

ABSTRACT

An implantable device for facilitating the healing of voids in bone, cartilage and soft tissue is disclosed. A preferred embodiment includes a cartilage region comprising a polyelectrolytic complex joined with a subchondral bone region. The cartilage region, of this embodiment, enhances the environment for chondrocytes to grow articular cartilage; while the subchondral bone region enhances the environment for cells which migrate into that region&#39;s macrostructure and which differentiate into osteoblasts. A hydrophobic barrier exists between the regions, of this embodiment. In one embodiment, the polyelectrolytic complex transforms to hydrogel, following the implant procedure.

CROSS REFERENCE

The present application is a continuation of U.S. patent applicationSer. No. 10/830,267, filed Apr. 21, 2004, which is a continuation ofU.S. patent application Ser. No. 10/199,961, filed Jul. 19, 2002, nowabandoned, which is a continuation-in-part of U.S. patent applicationSer. No. 09/909,027, filed Jul. 19, 2001, now abandoned, which is acontinuation-in-part of U.S. patent application Ser. No. 206,604, filedDec. 7, 1998, now U.S. Pat. No. 6,264,701, which is in turn a divisionof U.S. patent application Ser. No. 08/242,557, filed May 13, 1994. nowU.S. Pat. No. 5,981,825. The contents of each of the above-noted Patentsand applications is hereby fully incorporated herein by reference.

FIELD OF THE INVENTION

The present invention generally relates to the transport and/orculturing of cells, and more specifically to the healing of voids orother defects in bone, cartilage and soft tissue.

BACKGROUND OF THE INVENTION

The medical repair of bones and joints and other tissue in the humanbody presents significant difficulties, in part due to the materialsinvolved. Each bone has a hard, compact exterior surrounding a spongy,less dense interior. The long bones of the arms and legs, the thigh boneor femur, have an interior containing bone marrow. The material thatbones are mainly composed of is calcium, phosphorus, and the connectivetissue substance known as collagen.

Bones meet at joints of several different types. Movement of joints isenhanced by the smooth hyaline cartilage that covers the bone ends, bythe synovial membrane that covers the hyaline cartilage and by thesynovial fluid located between opposing articulating surfaces.

Cartilage damage produced by disease such as arthritis or trauma is amajor cause of physical deformity and dehabilitation. In medicine today,the primary therapy for loss of cartilage is replacement with aprosthetic material, such as silicone for cosmetic repairs, or metalalloys for joint realignment. The use of a prosthesis is commonlyassociated with the significant loss of underlying tissue and bonewithout recovery of the full function allowed by the original cartilage.The prosthesis is also a foreign body which may become an irritatingpresence in the tissues. Other long-term problems associated with thepermanent foreign body can include infection, erosion and instability.

The lack of a truly compatible, functional prosthesis subjectsindividuals who have lost noses or ears due to burns or trauma toadditional surgery involving carving a piece of cartilage out of a pieceof lower rib to approximate the necessary contours and inserting thecartilage piece into a pocket of skin in the area where the nose or earis missing.

Surgical removal of infected or malignant tissue is disfiguring and canhave harmful physiological and psychological effects. Regeneration ofsoft tissue, or tissue that mimics the natural properties of the removedtissue, can avoid or lessen these untoward consequences. Finally. adevice which delivers a therapy could aid the regeneration of tissue,minimize risk of infection, and/or treat any underlying disease orcondition.

The foregoing being exemplary, a device according to the teachings ofthe present invention is expected to add utility in many areas, seeTable 1, which is meant to be expansive of the foregoing, and notlimiting.

TABLE 1 Examples of tissues and procedures potentially benefiting fromthe teachings of the present invention Bone Bone tissue harvest Spinalarthrodesis Spinal fixation/fusion Osteotomy Bone biopsy Maxillofacialreconstruction Long bone fixation Compression fractures Hipreconstruction/replacement Knee reconstruction/replacement Handreconstruction Foot reconstruction Ankle reconstruction Wristreconstruction Elbow reconstruction Shoulder reconstruction CartilageMosaicplasty Meniscus Dental Ridge augmentation Third molar extractionTendon Ligament Skin Topical wound Burn treatment Biopsy Muscle DuraLung Liver Pancreas Gall bladder Kidney Nerves Artery Bypass SurgeryCardiac catheterization Heart Heart valve replacement Partial organremoval

In the past, bone has been replaced using actual segments of sterilizedbone or bone powder or porous surgical steel seeded with bone cellswhich were then implanted. In most cases, repair to injuries was madesurgically. Patients suffering from degeneration of cartilage had onlypain killers and anti-inflammatories for relief.

Until recently, the growth of new cartilage from either transplantationor autologous or allogeneic cartilage has been largely unsuccessful.Consider the example of a lesion extending through the cartilage intothe bone within the hip joint. Picture the lesion in the shape of atriangle with its base running parallel to the articular cavity,extending entirely through the hyaline cartilage of the head of thefemur, and ending at the apex of the lesion, a full inch (2.54 cm) intothe head of the femur bone.

Presently, there is a need to successfully insert an implant devicewhich will assure survival and proper future differentiation of cellsafter transplantation into the recipient tissue defect. Difficultieshave been experienced with engineering the implant environment such thatcells may survive, and also with supporting proper cell differentiation.

Presently, for example, cartilage cells, called chondrocytes, whenimplanted along with bone cells, can degenerate or dedifferentiate intomore bone cells. Because hyaline cartilage is an avascular tissue, itmust be protected from intimate contact with sources of high oxygentension such as blood. Bone cells, in contrast, require high oxygenlevels and blood. For this reason, the subchondral bone region of thedevice should be isolated from the cartilage region, at least so far asoxygen and blood are concerned.

Most recently, two different approaches to treating articular lesionshave been advanced. One approach such as disclosed in U.S. Pat. No.5,041,138 is coating bioderesorbable polymer fibers of a structure withchemotactic ground substances. No detached microstructure is used. Theother approach such as disclosed in U.S. Pat. No. 5,133,755 useschemotactic ground substances as a microstructure located in voids of amacrostructure and carried by and separate from the biodegradablepolymer forming the macrostructure. Thus, the final spatial relationshipof these chemotactic ground substances with respect to the bioresorbablepolymeric structure is very different in U.S. Pat. No. 5,041,138 fromthat taught in U.S. Pat. No. 5,133,755.

The fundamental distinction between these two approaches presents threedifferent design and engineering consequences. First, the relationshipof the chemotactic ground substance with the bioresorbable polymericstructure differs between the two approaches. Second, the location ofbiologic modifiers carried by the device with respect to the device'sconstituent materials differs. Third, the initial location of theparenchymal cells differs.

Both approaches employ a bioresorbable polymeric structure and usechemotactic ground substances. However, three differences between thetwo approaches are as follows.

I. Relationship of Chemotactic Ground Substances with the BioresorbablePolymeric Structure

The design and engineering consequence of coating the polymer fiberswith a chemotactic ground substance is that both materials become fusedtogether to form a single unit from structural and spatial points ofview. The spaces between the fibers of the polymer structure remaindevoid of any material until after the cell culture substances areadded.

In contrast, the microstructure approach uses chemotactic groundsubstances and/or other materials, separate and distinct from themacrostructure. The microstructure resides within the void spaces of themacrostructure. Additionally, an embodiment incorporating amicrostructure may use materials such as polysaccharides and chemotacticground substances that are spatially separate from the macrostructurepolymer thereby forming an identifiable microstructure, separate anddistinct from the macrostructure polymer.

The design and engineering advantage to having a separate and distinctmicrostructure capable of carrying other biologically active agents canbe appreciated in the medical treatment of articular cartilage. RGDattachment moiety of fibronectin is a desirable substance for attachingchondrocytes cells to the lesion. However, RGD attachment moiety offibronectin is not, by itself, capable of forming a microstructure ofvelour in the microstructure approach. Instead, RGD may be blended witha microstructure material prior to investment within macrostructureinterstices.

II. Location of Biologic Modifiers Carried by a Device with Respect tothe Device's Constituent Materials

Coating only the polymer structure with chemotactic ground substancesnecessarily means that the location of the chemotactic ground substanceis only found on the macrostructure (e.g., bioresorbable polymer)fibers, thereby affording a two dimensional presentation. Themicrostructure approach uses the microstructure to carry biologicmodifiers (e.g., growth factors, morphogens, drugs, etc.), however thepresentation is analogous to a three dimensional presentation.Therefore, the coating approach has a limited capacity to carry biologicmodifiers with the biodegradable polymeric structure.

III. Initial Location of the Parenchymat Cell

Because the coating approach attaches the chemotactic ground substancesto the surfaces of the structure and has no microstructure resident inthe void volume of the device, the coating approach precludes thepossibility of establishing a network of extracellular matrix material,specifically a microstructure, within the spaces between the fibers ofthe polymer structure once the device is fully saturated with cellculture medium. The coating approach predetermines that any cellsintroduced via culture medium will be immediately attracted to thesurface of the structure polymer and attach thereto by virtue of thechemotactic ground substances on the polymer's surfaces.

The consequence of confining chemotactic ground substances to only thesurfaces of the polymeric structure places severe restrictions on thenumber of cells that can be accommodated by the coated device.

In contrast with the coating approach, the microstructure approach, bylocating chemotactic ground substances in the void spaces of the device,makes available the entire void volume of the device to accommodate theattracted cells which then lay down their own extracellular matrixresulting in a more rapid and complete tissue regrowth or ingrowth.

One of the many objects of this invention, as will be discussed, is toprotect and aid cellular ingrowth or regeneration of various types ofnew tissue, as well as providing methods of concurrent delivery oftherapies and other treatments.

SUMMARY OF THE INVENTION

A device of the present invention is a prosthesis or implant for in vivoculturing of tissue cells in a diverse tissue or homogeneous lesion. Theentire macrostructure, or a major portion, of this device may becomposed of a bioresorbable polymer. Alternatively, the microstructuremay be the only portion of the device which is resorbable, if amicrostructure was employed at all. Alternatively, it is also conceivedthat the device could be used to culture cells via in vitro techniquesknown in the art for later in vivo transplantation.

A device of the present invention may include a macrostructure,microstructure, free precursor cells cultured in vitro or from tissue,or biologically active agents. “Biologically active agents” as used inthis disclosure meaning, but not limited to, growth factors, morphogens,drugs, proteins, cells, cellular components. signaling proteins, signaltransduction factors, and other therapeutic agents.

An anatomically specific device of the present invention could bedesigned primarily for treating cartilage and bone lesions and, whenused for that purpose, preferentially has two main regions: a cartilageregion and a subchondral bone region. Alternatively, it is envisionedthat a singular region may be employed to repair defects in other areasand types of host tissue. Likewise, additional regions may be used to“bridge” tissue of distinct histological variation, as well as othervariations.

A first embodiment of the present invention comprises a cartilage regionwhich has a macrostructure and a microstructure. The selectiveconcentration gradient of material in the microstructure may beselectively varied within certain regions of the macrostructure voids toaffect different biologic characteristics and tissue requirements.

The microstructure of a single device of the present invention may becomposed of multiple different materials, some without chemotacticproperties, in different regions of macrostructure void space dependingupon varying tissue and biologic characteristics and requirements.

The subchondral bone region of this embodiment includes a macrostructurecomposed of a biologically acceptable, polymer (preferablybioresorbable) arranged as a one piece porous body with “enclosedrandomly sized, randomly positioned and randomly shaped interconnectingvoids, each void communicating with all the others, and communicatingwith substantially the entire exterior of the body” (quoted portion fromU.S. Pat. No. 4,186,448). In the preferred embodiment as described here,the internal three dimensional architecture of the macrostructureresembles that of cancellous bone. In other embodiments, the internal3-D architecture of the macrostructure may be highly ordered, asdescribed in U.S. Pat. No. 5,981,825, to replicate the spatial patternsof other tissues or to create a tissue pattern required for performanceof specific anatomic and/or physiologic functions. In one preferredembodiment, polylactic acid (PLA), fabricated in the 3-D architecture ofintercommunicating voids described above forms the macrostructure. Othermembers of the hydroxy acid group of compounds can also be used as canany bioresorbable polymer, natural or synthetic, if fabricated into asimilar architecture. Alternatively, the macrostructure could befabricated from natural materials (e.g., bone, coral, or collagen),ceramic materials (whether natural or synthesized, e.g., hydroxyapatiteor tricalcium phosphate), or other materials, such as those shown inTables 2 and 3.

The gross, or macro, structure of this embodiment attempts to addressthree major functions for chondrogenesis and osteogenesis: 1) restoresmechanical architectural and structural competence; 2) providesbiologically acceptable and mechanically stable surface structuresuitable for genesis, growth and development of new non-calcified andcalcified tissue; and 3) functions as a carrier for other constituentsof the present invention which do not have mechanical and structuralcompetence.

The microstructure of this embodiment may be composed of variouspolysaccharides which, in a preferred form, is alginate but can also behyaluronic acid (abbreviated by HY). Interstices of the polylactic acidmacrostructure of the body member are invested with the microstructuresubstance which may be in the form of a velour having the samearchitecture of interconnecting voids as described for themacrostructure, but on a microscopic scale. Functions of themicrostructure (e.g., HY) may include: 1) attraction of fluid bloodthroughout the device; 2) chemotaxis for mesenchymal cell migration andaggregation; 3) carrier for osteoinductive and chondro-inductiveagent(s); 4) generation and maintenance of an electro-negative woundenvironment; 5) agglutination of other connective tissue substances witheach other and with itself, and 6) coating of the edges of themacrostructure to minimize or prevent foreign body giant cell responses,as well as other adverse responses to the implant. Other examples ofsuitable microstructures are fibronectin and, especially for thereconstruction of articular cartilage, an RGD attachment moiety offibronectin.

The osteoinductive agent, bone morphogenetic protein, has the capacityto induce primitive mesenchymal cells to differentiate into bone formingcells. Another osteogenic agent, bone derived growth factor, stimulatesactivity of more mature mesenchymal cells to form new bone tissue. Otherbiologically active agents which can be utilized, especially for thereconstruction of articular cartilage, include but are not limited totransforming growth factor beta (TGF beta) and basic fibroblast growthfactor (bFGF).

In this first embodiment, as well as the balance of the specificationand claims, the term “bioabsorbable” is frequently used. There existssome discussion among those skilled in the art, as to the precisemeaning and function of bioabsorbable material (e.g., polymers), and howthey differ from resorbable, absorbable, bioresorbable, biodegradable,and bioerodable materials. The current disclosure contemplates all ofthese materials, and combines them all as bioresorbable. Any use of analternate disclosed in this paragraph is also meant to describe andinclude all of the others.

In a second embodiment of the present invention, the device acts as atransport device for precursor cells harvested for the production ofconnective tissue. The device can be press fit into the site of lesionrepair, and subsequently charged with a solution of cells, growthfactors, etc., as will be described later. Another aspect of thisembodiment is that the microstructure velour can be treated with an RGDattachment moiety of fibronectin that facilitates the attachment of freeprecursor cells to be carried to the lesion repair site.

Additional embodiments of the present invention allow for the tailoringof mechanical and physical properties through the use of additions ofother polymers, ceramics, microstructures and processes (e.g., voidtailoring, cross-linking, and pre-stressing). Additionally, the deliveryof therapies aids regeneration of tissue, minimizes proceduraldiscomfort to the patient, and treats underlying disease.

DESCRIPTION OF THE PREFERRED EMBODIMENTS

A device and methods according to the preferred teachings of the presentinvention are disclosed for treating mammalian bone and cartilage andsoft tissue deficiencies, defects, voids and conformationaldiscontinuities produced by congenital deformities, osseous and/or softtissue pathology, traumatic injuries, and accidental, surgical, orfunctional atrophy. The primary purpose of this implant device is toprovide the means by which chondrocytes, or other cells, and theirattendant synthesis, cultured in vitro, can be transported into a defectand be safely established therein. Thus, the most preferred embodimentsof the present invention provides means to regenerate a specific form oftissue.

A first embodiment of the present invention consists of two main parts,the cartilage region and the subchondral bone region joined at aninterface surface. Each of the cartilage and the subchondral boneregions of the device includes a macrostructure composed of abioresorbable polymer either as homogeneous polymers or combinations oftwo or more co-polymers from groups of, for example, poly (alpha-hydroxyacids), such as polylactic acid or polyglycolic acid or theirco-polymers, polyanhydrides, polydepsipeptides, or polyorthoester.Devices fabricated for prototypes of animal studies to-date have beenfabricated from the homopolymer D, D-L, L-polylactic acid, andpolyelectrolytic complexes.

The bioresorbable polymer in the subchondral bone region in this form isin the architecture of cancellous bone such as of the type described inU.S. Pat. Nos. 4,186,448 and 5,133,755, which are hereby incorporatedherein by reference.

The architecture of the cartilage region may be formed utilizingestablished techniques widely practiced by those skilled in the art ofbioresorbable polymers. These methods include injection molding, vacuumfoaming, spinning hollow filaments, solvent evaporation, solubleparticulate leaching or combinations thereof. For some methods,plasticizers may be required to reduce the glass transition temperatureto low enough levels so that polymer flow will occur withoutdecomposition.

The macrostructure polymer of the cartilage region is joined or bound tothe macrostructure polymer of the subchondral bone region by a processsuch as heat fusion which does not involve the use of solvents orchemical reactions between the two polymer segments. The resulting unionbetween the two architectural regions is very strong and can withstandany handling required to package the device as well as any forcesdelivered to it as a result of the implantation technique withoutdistorting the device's internal architecture of void spaces.

In former constructs such as U.S. Pat. No. 5,133,755, the preferredmicrostructure was hyaluronan which is synonymous with hyaluronic acid,hyaluronate, HA and HY. The hyaluronan was distributed uniformlythroughout the internal void volume of the device. According to theteachings of the present invention, an option is provided of selectingwhether or not the microstructure, if any, should be dispersedthroughout all the void spaces depending on whether the arrangement isbeneficial to the tissues being treated. A device of the presentinvention permits incomplete dispersal as desired or complete dispersalthroughout the entire void volume of the device but expressingconcentration gradients of microstructure material as a means ofcontrolling transplanted cell population numbers within the device'sinternal domains.

A dry filamentous velour of chemotactic ground substance, for exampleRGD attachment moiety of fibronectin carried by hyaluronic acid oralginic acid velour, may be established within the void spaces of thedevice. Upon saturation with water, water-based cell culture media orfluid blood, the dry velour of chemotactic ground substance is dissolvedinto a highly viscous gel which maintains the chemotactic groundsubstance as a network of dissolved polysaccharide strands, stillsuspended within the void volume of the polymeric macrostructure. It isenvisioned that other therapies may also be carried by this gel, as willbe discussed later.

If the cell culture media is a fluid which saturates the device andcreates the gel, then those cells suspended in the culture medium willbe temporarily trapped within the gel due to the gel viscosity. Thedegree of gel viscosity and the length of time the gel maintainssignificantly high viscosities will aid in cellular propagation, i.e.,restraining the transported cells by means of microstructure gel givesthe cells additional time to execute biological processes. Additionally,this restraint can be used to modulate the delivery rate of the therapy.

The volume of space once occupied by the microstructure gel can then beoccupied by the interstitial fluid and increased numbers of cells. Inthe articular cartilage regeneration of the preferred form, it isdesired to protect the transplanted cells from access to fluid blood andcollateral circulation. In other tissue regeneration situations,however, it may be desirable or beneficial to attract fluid blood intothe device's interstices as quickly as possible. In these situations,therefore, fibrin (i.e. blood clot), endothelial cells, or othermaterials or therapies may be loaded into the device, or gained fromsources of viable collateral circulation.

Certain embodiments of the present invention depart from prior practiceby strategically positioning the microstructure material in thatspecific portion of the device which performs particular functionsunique to the mature anatomy being regenerated in that vicinity. Suchsegregation of microstructure material within the device is based on theneed to endow one portion of the device with special biologic functionsthat must be isolated from the remainder of the implanted device.

In yet another embodiment of the present invention, the microstructurehas a secondary purpose to present enough chondrocytes to thesubehondral bone region immediately adjacent to the cartilage region toinsure that a competent osteo-chondral bond is established between thenewly developed cartilage and the newly developed bone.

Within the inventive concept of several embodiments of the presentinvention is the establishment of variations in the concentration ofmicrostructure within the void space network of the macrostructure inorder to assure that the therapeutic elements and biologically activeagents brought from in vitro culture, or loaded as will be describedlater, are present within the final device in greatest quantity wherethey are most needed. Such variations in concentration can beaccomplished by varying concentrations of microstructure solutions priorto investment into macrostructure voids of the device or regions thereofbefore joining, as well as other methods known in the art.

In yet another embodiment of the current invention, the cartilage regionof the construct comprises a polyelectrolytic complex (PEC). Thiscomplex preferably comprises polyanions and polycations. Since certainof these complexes in their dry states may not have sufficient strengthto allow handling, processing may be required to increase theirstructural integrity. This processing can follow the methods previouslydisclosed, as well as various other generic techniques known to thoseskilled in the art. Because of the unique bonding structures containedin PEC's, some researchers have referred to them as poly-ionic complexes(PIC's). For this reason, the current disclosure recognizes nodifference between the PEC and the PIC.

The PEC may be formed from glycosaminoglycans (GAG's) and polycations aswell as other similarly structured compounds. While having the requisiteelectron affinity noted above for bonding, some of the sulfonated GAG'smay not be effective in attracting the appropriate cell-types. In apreferred embodiment, the PEC is made from hyaluronic acid (HY), anon-sulfonated GAG, and chitosan. The PEC may be fabricated by variousmethods known to those skilled in the art, one such method follows.

The strong negative charge associated with HY is provided by thecarboxylic acid group (—COOH) of its glucuronic acid moiety. Whenexposed to pH levels below about 6.5, the amine groups of chitosanmolecules become protonated, thus rendering the molecules soluble inwater and providing them with a strong positive charge that attractsnegatively-charged molecules (e.g., HY, etc.) and thus formingelectrostatic interactions. When a solution of protonated chitosan isexposed to a solution of HY, an insoluble precipitate (the PEC) isformed.

In yet another PEC embodiment, the PEC is made from hyaluronic acid andcollagen (i.e., collagen type I or type II or type III, etc.), wherecollagen acts as a polycation. Collagen, an amphoteric species,functions as a cation when treated similarly to chitosan, as describedabove, or by other methods known to those skilled in the art.

The collagen may be supplied to the PEC in the form of demineralizedbone matrix (DBM) material. It is realized that DBM also comprises, inaddition to collagen, morphogens and growth factors, as secondaryconstituents. It is also recognized that these secondary constituentsmay add to the overall tissue regenerative capacity of the implant.

Other glycosaminoglycans such as, but not limited to, heparin,chondroitin-4-SO.sub.4, chondroitin-6-SO.sub.4, dermatan-SO.sub.4, andkeratin sulfate may also be used as a complement to or in place ofhyaluronic acid, in these various embodiments.

In a similar embodiment, the macrostructure or microstructure, if any,of any region(s) may comprise chitosan, not bound in the aforementionedPEC. This embodiment, herein referred to as a “regeneration complex” maybe formed by the techniques discussed herein, as well as those othersknown in the art. Alternatively, this regeneration complex may comprisea protein (e.g., type I collagen, type II collagen, type III collagen,carrageenan, fibrin, elastin, resilin, abductin, demineralized bone, oragarose), polysaccharide (e.g., cellulose, starches, chitosan, alginate,sulfated glycosaminoglycans, or non-sulfated glycosaminoglycans), alipid (e.g., phospholipid, triglyceride, waxes, steroids,prostaglandins, or terpenes), a synthetic polymer (e.g., polylactide,polyglycolide, polyurethane, polyethylene, poly-e-caprolactone,polyvinyl alcohol, polycarbonate, or PTFE), ceramic (e.g., bioglass orcalcium phosphate), singularly or as a mixture thereof. Thesealternatives may be formed by methods similar to those used formonolithic chitosan, as well as those previously disclosed.

By way of example, one embodiment utilizes a resorbable polymermacrostructure and hyaluronic acid microstructure in one region that isadjacent to a collagen regeneration complex. The collagen can be ofseveral varieties as well as composites of thereof. Kensey NashCorporation (Exton, Pa.) manufactures soluble collagen known as Semed S,fibrous collagen known as Semed F, and a composite collagen known asP1076. Each of these materials would be suitable for this embodiment.This embodiment may also include additives (e.g., sodium hyaluronate)blended or composited with the collagen slurry and co-lyophilized tocreate a material with desirable mechanical and chemical properties. Theregeneration complex may undergo chemical, thermal, or radiationtreatments in order to cross-link the material to provide desiredstrength and/or degradation qualities. Additionally, a calcium mineralsuch as hydroxyapatite or a growth factor, such as TGF-beta, may beadded to the regeneration complex or to the neighboring region(s) inorder to customize the implant for use in a bone or cartilageregeneration device. All of the foregoing alterations of the device'smechanical, chemical, or biological properties and responses arereferred to as “matrix matching.”

Matrix matching may also be achieved by processes other thancross-linking. For example, pore size, shape, and population may beengineered, by degree and rate of lyophilization, the polymer structuremay be plastically strained or directionally treated to impartanisotropy or the like. As has been described, macrostructure andmicrostructural additions can greatly affect the degree of matrixmatching; not only by the properties of addition (i.e., relative to theproperties of the host matrix), but also by the relative amount placedtherein (i.e., relative to total amount of macrostructure, or totalamount of void space available to be filled by the microstructure).

Such matrix matching may be employed to approximate or nearlyapproximate the property of the host or other desired tissue to beregenerated. Alternatively, where the aforementioned result is notfeasible, desirable (e.g., due to patient discomfort, allowances forinflammation of existing tissue, or sacrificing some strength for addedtoughness), or practical, the degree of matrix matching may beintentionally limited. While several exemplary embodiments have beengiven, additional composite elements and additives are contemplated(e.g., including PEC complexes and regeneration complexes, andcombinations thereof), many of which are listed in Tables 2 and 3.Various other processes are also known in the art, which may be usedalone, or in combination with any of the foregoing, in order toaccomplish this same effect and result.

The tissue resulting after ingrowth or regeneration may also be matrixmatched, that is, the tissue strength, density, and pliability may bealtered by the matrix used. Ideally, the device would be matrix matched,and so would the regenerated tissue, although matrix matching refers toeither, as is discussed in more detail later.

Another similar embodiment utilizes a demineralized bone matrixmacrostructure and hyaluronic acid microstructure in one region that isadjacent to a chitosan PEC, as is described above, on a first side and achitosan PEC on a second diametrically opposed side. This multi-layeredimplant would have the ability to regenerate cancellous bone through itsmiddle region while regenerating cortical bone or cartilage on the endregions. Bone or cartilage are used in this example, but various othertissues are contemplated, and the regions may be arranged other thanuniaxially. Additionally, an embodiment is contemplated wherein thedemineralized bone may be replaced with porous hydroxyapatite if astronger implant or longer-lasting type implant is desired.

In yet another embodiment, collagen may be used, for example, in theform of a porous fabric, to define a macrostructure. The porous fabriccan be created to allow for specific pore size and separation. Thefabric maintains an architecture that is suitable and similar to theatmosphere that chondrocytes are exposed to in host tissue. Thismacrostructure presents the structural integrity necessary to supply ahomeostatic atmosphere for chondrocyte viability. This allowsregenerative cascades to occur and allows for replication of damagedtissue. In addition, elements may be added to the macrostructure tocreate one microstructure. An example of this can be hyaluronic acid,demineralized bone matrix DBM, etc. Regardless of whether or not amicrostructure is used, the macrostructure region may be attached to asecond region via a porous polymeric film.

This film may be interposed between the first (e.g., collagen) andsecond regions at their interface, thereby increasing the strength ofthe bond. This interposition may be formed in a manner similar to thefollowing example; a porous or non-porous film may be created of thedesired polymer to create the needed bond. The thin film may be placedbetween the two regions to allow fixation in such manner where heat, UV,etc. may be used to combine the two materials.

Additionally, the polymer film may be constructed with the use of asolvent to create the film. This solution/slurry/suspension/gel emulsioncan be applied to both or either material, with varying concentrationsto bind the two materials. An example of one such procedure would be toapply the solution in such a fashion where a brush would be used forapplication. By way of example and not limitation, other manners may beemployed, including spraying, dipping, etc. Therefore, these embodimentsdescribe the application of the film in the liquid and/or solid states,and this disclosure contemplates other methods of polymer depositionknown to those skilled in the art (e.g., spraying, dipping, heatapplication, UV, etc.)

In the foregoing PEC and regeneration complex embodiments, it is alsocontemplated that these devices will be implanted into a tissuerequiring regeneration of one or multiple tissues. The devices of thisdisclosure may be implanted in a variety of ways. In one embodiment, theimplant will be pressed into a defect site and, as will be discussedlater in greater detail, will expand in apparent volume thus maintainingpositive contact with host tissue. Other methods of implantation includesuturing the implant into place, suturing a flap over the implant (suchas a periosteal flap), using a glue or sealant (such as a fibrin glue),screws and fixturing, containing the implant within a separate devicewhich is screwed, glued (e.g. thrombin, cyanoacrylate, etc.),press-fitting in place (such as an interbody fusion cage), or by othermethods known to those skilled in the art.

Additionally, the shape or contouring of the implant can be used to holdthe implant in place. In one embodiment, the implant may be created inthe shape of a screw or a barb by using a mold, by cutting away thematerial, or by other methods known to those skilled in the art. Inanother embodiment, the contouring is created only in a region of theimplant where tissue will regenerate the fastest. The contouring ispurposely designed to provide resistance to shear, tensile, compressive,torque, and other forces acting to dislodge the implant. While someapplications require contouring in only one region, other applicationswill require multiple regions of contouring.

The foregoing PEC and regeneration complex embodiments will have certainbeneficial reactions following implant. That is, among other things,particular of these formations will imbibe water-based fluids in theimplant vicinity. This fluid infusion will cause one or more regions ofthe implant to swell. Swelling may be important for securement reasons,as previously discussed, or for its affect on biological activity.

The swelling of these particular implant compositions is nearlyequiaxial, that is, proportional in all directions to dimensions of theoriginal, dry construct. Upon prolonged hydration, void spaces of thedry construct become occluded by the gel generated when water becomesbound to fibers of the PEC and additional water becomes entrappedbetween hydrated PEC filaments. Thus, those skilled in the art refer tothis resulting structure as a hydrogel. The hydrogel medium endows itsregion of the device with several benefits that include, but are notlimited to: (i) restricting trans-implant communication of biologicallyactive agents; (ii) allowing its cargo of biologically active agentsunrestricted access to host tissues immediately after implantation whileprogressively restricting this access over time; (iii) providing a depotof biologically active agent for access by cells entering the hydrogelregion; and (iv) establishing the early microenvironment for cellmigration into the defect (e.g., chemotaxis). The minimization of theaccess to these agents, however, is not detrimental to the function ofthe implant, since mass transfer (i.e., transfer of gases, nutrients,and cell waste products) occurs through hydrogels, and the cellularfunctions of respiration and metabolism continue.

In the foregoing PEC and regeneration complex embodiments, it iscontemplated that the subehondral bone region comprises a resorbablepolymer (polymer being synthetic or organic/natural, e.g., see Table 2)as well as other non-resorbable or non-polymeric materials (e.g., seeTable 3); additionally, these materials may be used for a PEC regionmacrostructure, if one is employed. The macrostructure being a structurecomprising voids, in which the PEC could be invested, along with othermaterials and therapies. In this type of embodiment, the materials andtherapies are referred to collectively as the microstructure. Themacrostructure and microstructure are also tailorable by other additions(e.g., see those materials and compounds listed in Tables 2 and 3).

In another embodiment, the void spaces within the macrostructure ormicrostructure, of any region, cause cellular regeneration effects bythe size and/or shape thereof. That is, the relative size of the voidspace can affect the resulting cellular structure that is generated, orlikewise the shape of the void space can affect cellular structure.Thus, engineering the size or shape of void spaces to stress orconstrict cellular function can influence forms of regenerated tissue.

Similarly, the mechanical properties (e.g., density, hardness, modulusof elasticity, or compressive stiffness) or physical properties (e.g.,macrostructure void, microstructure or a void therein, or cellattachment aiding material which is in the microstructure) of the hoststructure can alter the cellular reproduction type or phenotype. This isexpected to be caused by the interaction between the host material andthe endocellular fibrils, but other actions and reactions areanticipated to contribute to this effect. This interaction may beutilized to tailor the resulting cell type, by tailoring the hostmaterial's mechanical or physical properties.

TABLE 2 Examples and Sub-types of Bioresorbable Polymers forConstruction of the Device Macrostructure and/or Microstructure of theCurrent Invention Aliphatic polyesters Bioglass Cellulose ChitinCollagen Types 1 to 20 Native fibrous Soluble Reconstituted fibrousRecombinant derived Copolymers of glycolide Copolymers of lactideElastin Fibrin Glycolide/l-lactide copolymers (PGA/PLLA)Glycolide/trimethylene carbonate copolymers (PGA/TMC) HydrogelLactide/tetramethylglycolide copolymers Lactide/trimethylene carbonatecopolymers Lactide/.epsilon.-caprolactone copolymersLactide/.sigma.-valerolactone copolymers L-lactide/dl-lactide copolymersMethyl methacrylate-N-vinyl pyrrolidone copolymers Modified proteinsNylon-2 PHBA/.gamma.-hydroxyvalerate copolymers (PHBA/HVA)PLA/polyethylene oxide copolymers PLA-polyethylene oxide (PELA) Poly(amino acids) Poly (trimethylene carbonates) Poly hydroxyalkanoatepolymers (PHA) Poly(alklyene oxalates) Poly(butylene diglycolate)Poly(hydroxy butyrate) (PHB) Poly(n-vinyl pyrrolidone) Poly(orthoesters) Polyalkyl-2-cyanoacrylates Polyanhydrides PolycyanoacrylatesPolydepsipeptides Polydihydropyrans Poly-dl-lactide (PDLLA)Polyesteramides Polyesters of oxalic acid Polyglycolide (PGA)Polyiminocarbonates Polylactides (PLA) Poly-l-lactide (PLLA)Polyorthoesters Poly-p-dioxanone (PDO) Polypeptides PolyphosphazenesPolysaccharides Polyurethanes (PU) Polyvinyl alcohol (PVA)Poly-.beta.-hydroxypropionate (PHPA) Poly-.beta.-hydroxybutyrate (PBA)Poly-.sigma.-valerolact-one Poly-.beta.-alkanoic acids Poly-.beta.-malicacid (PMLA) Poly-.epsilon.-caprolactone (PCL) Pseudo-Poly(Amino Acids)Starch Trimethylene carbonate (TMC) Tyrosine based polymers

TABLE 3 Examples of alternative materials that may be used for themacrostructure and/or microstructure of the current invention AlginateBone allograft or autograft Bone Chips Calcium Calcium Phosphate CalciumSulfate Ceramics Chitosan Cyanoacrylate Collagen Dacron Demineralizedbone Elastin Fibrin Gelatin Glass Gold Glycosaminoglycans HydrogelsHydroxy apatite Hydroxyethyl methacrylate Hyaluronic Acid LiposomesMesenchymal cells Nitinol Osteoblasts Oxidized regenerated cellulosePhosphate glasses Polyethylene glycol Polyester PolysaccharidesPolyvinyl alcohol Platelets, blood cells Radiopacifiers Salts SiliconeSilk Steel (e.g. Stainless Steel) Synthetic polymers Thrombin TitaniumTricalcium phosphate

It is also contemplated that the PEC region, or the regeneration complexregion, may be used alone (i.e., without a subchondral bone, or otherregion) or with a microstructure contained therein. Furthermore, it isrecognized that when two or more regions are joined, as discussed in thevarious embodiments herein, there may exist a zone that is chemically orstructurally distinct from either of, or one of, the regions. This maybe incidental to the processing methods employed, or the naturalreaction of the body's incorporation of the implant. That is, the zonemay be intentional or a planned or unplanned result. For example, zonesincorporating barriers and other active agents are within the scope ofthe invention. Furthermore, a zone incorporating a hydrophobic barrierwherein the surface properties of the macrostructure are altered (e.g.,rendered hydrophobic) without altering the geometry or mechanicalcharacteristics of the macrostructure is envisioned.

It is further contemplated that gene therapy may be used with PECconstructs, or similar devices for the regeneration of bone and softtissue. Gene therapies are currently of two primary types, and are bothtogether hereinafter referred to as “gene therapy” or “engineeredcells”. However, others are anticipated. The primary methodologies andbasic understandings are described herein (see also Table 4).

First, nucleic acids may be used to alter the metabolic functioning ofcells, without altering the cell's genome. This technique does not alterthe genomic expressions, but rather the cellular metabolic function orrate of expression (e.g., protein synthesis).

Second, gene expression within the host cell may be altered by thedelivery of signal transduction pathway molecules.

In a preferred embodiment, mesenchymal stem cells are harvested from thepatient, and infected with vectors. Currently, preferred vectors includephages or viri (e.g., retrovirus or adenovirus). This preferredinfection will result in a genetically engineered cell, which may beengineered to produce a growth factor (e.g., insulin like growth factor(IGF-1)) or a morphogen (e.g., bone morphogenic protein (BMP-7)), etc.(see also those listed in Table 4). Methods of infection as well asspecific vectors are well known to those skilled in the art, andadditional ones are anticipated. Following this procedure, thegenetically engineered cells are loaded into the implant. Cytokines asdescribed and used herein are considered to include growth factors.

Loading of the cells in this embodiment may be achieved prior to,during, or immediately following the implantation procedure. Loading maybe achieved by various methods including, but not limited to, byinjecting a solution containing the engineered cells into the implant,by combining the cells with the macrostructure, or by any void fillingcomponent, or by themselves, in the void spaces of any of the regions.Prior to the loading of fluid, whether by manual injection or byinfiltration from the implant site, the PEC is referred to as being in a“dry state.”

Other therapies, including but not limited to drugs, biologically activeagents, and other agents, may also be utilized in or with the PEC, orany other associated or adjoined region (e.g., macrostructure ormicrostructure); either to aid the function of the PEC and/or any otherassociated or adjoined region or to cause other stimuli. The drugs,biologics, or other agents may be naturally derived or otherwise created(e.g. synthesized). For example, growth factors can be derived from aliving being (e.g. autologous, bovine derived, etc.), producedsynthetically, or made using recombinant techniques (e.g. rhBMP-2).Regardless of the time of investment or incorporation of thesematerials, they may be in solid particulate, solution gel or otherdeliverable form. Utilizing gel carriers may allow for the materials tobe contained after wetting, for some tailorable length of time.Furthermore, additions may be incorporated into the macrostructureduring manufacture or later. The incorporations may be made by blendingor mixing the additive into the macrostructure or microstructurematerial, by injection into the gel or solid material, or by othermethods known to those skilled in the art. Another method ofincorporating additives, biologics and other therapies, into themacrostructure or microstructure of one or more regions of the device isthrough the use of micro spheres.

The term “microsphere” is used herein to indicate a small additive thatis about an order of magnitude smaller (as an approximate maximumrelative size) than the implant. The term does not denote any particularshape. It is recognized that perfect spheres are not easily produced.The present invention contemplates elongated spheres and irregularlyshaped bodies.

Microspheres can be made of a variety of materials such as polymers,silicone and metals. Biodegradable polymers are ideal for use increating microspheres (e.g., see those listed in Tables 2 and 3). Therelease of agents from bioresorbable microparticles is dependent upondiffusion through the microsphere polymer, polymer degradation and themicrosphere structure. Although most any biocompatible polymer could beadapted for this invention, the preferred material would exhibit in vivodegradation. It is well known that there can be different mechanismsinvolved in implant degradation like hydrolysis, enzyme mediateddegradation, and bulk or surface erosion. These mechanisms can alone orcombined influence the host response by determining the amount andcharacter of the degradation product that is released from the implant.The most predominant mechanism of in vivo degradation of syntheticbiomedical polymers like polyesters, polyamides and polyurethanes, isgenerally considered to be hydrolysis, resulting in ester bond scissionand chain disruption. In the extracellular fluids of the living tissue,the accessibility of water to the hydrolysable chemical bonds makeshydrophilic polymers (i.e. polymers that take up significant amounts ofwater) susceptible to hydrolytic cleavage or bulk erosion. Severalvariables can influence the mechanism and kinetics of polymerdegradation, including but not limited to material properties likecrystallinity, molecular weight, additives, polymer surface morphology,and environmental conditions. As such, to the extent that each of thesecharacteristics can be adjusted or modified, the performance of thisinvention can be altered.

In a homogeneous embodiment (i.e., monolithic or composite of uniformheterogeneity) of a therapy delivering implant material, the deviceprovides continuous release of the therapy over all or some of thedegradation period of the device. In an embodiment incorporatingmicrospheres, the therapy is released at a preferential rate independentof the rate of degradation of the matrix resorption or degradation. Incertain applications, it may also be necessary to provide a burstrelease or a delayed release of the active agent. The device may also bedesigned to deliver more than one agent at differing intervals anddosages. This time-staged delivery also allows for a dwell ofnon-delivery (i.e., a portion not containing any therapy), therebyallowing alternating delivery of non-compatible therapies. Deliveryrates may be affected by the amount of therapeutic material, relative tothe amount of resorbing structure, or the rate of the resorption of thestructure.

Time-staged delivery may be accomplished via microspheres, in a numberof different ways. The concentration of therapeutic agent may varyradially, that is, there may be areas with less agent, or there may beareas with no agent. Additionally, the agent could be varied radially,such that one therapy is delivered prior to a second therapy allowingthe delivery of noncompatible agents, with the same type of sphere,during the same implant procedure. The spheres could also vary incomposition. That is, some portion of the sphere population couldcontain one agent, while the balance may contain one or more alternateagents. These differing spheres may have different delivery rates.Finally, as in the preceding example, there could be different deliveryrates, but the agent could be the same, thereby allowing a burst dosefollowed by a slower maintained dose.

In a time-phased delivery embodiment, the implant may be constructed toeffect a tailored delivery of active ingredients. Both the presence ofthe implant and the delivery of the select agents are designed to leadto improvements in patients with tissue defects, as a result ofdelivering in no certain order: (1) a substratum onto which cells canproliferate, (2) a drug or biologically active agent which can act as asignaling molecule which can activate a proliferating or differentiatingpathway, (3) a drug or biologically active agent which may act as adepot for nutrients for proliferating and growing cells, and (4) a drugor biologically active agent which will prevent an adverse tissueresponse to the implant, or provide a therapy which reduces infectionand/or treats an underlying disease or condition.

In yet another embodiment, a matrix matched device is designed to mimicthe properties of the host tissue and/or shape of any removed tissue,immediately upon implant or shortly after absorbing bodily fluids intothe device's void network, or microstructure (if one is employed). Thechanging properties of certain polymers, following absorption oradsorption, of fluids is well known in the art. The device will afford amore natural feeling (than traditional implants), and minimize thefeeling of a foreign body to the patient. As the device resorbs, it willfoster the ingrowth or regeneration of tissue with properties matchingor nearly approximating the host tissue, such that after a certainperiod of time (e.g., about two months to two years), the site of theprocedure may have the pre-procedure look and feel restored. Thisembodiment may be especially beneficial for patients who have organs,tumors, or other tissue masses removed, and affords all of thetherapeutic modes of the previous embodiments.

The device may matrix match the resulting tissue by preferentiallyaltering the resulting scar tissue that is developed. Normal scar tissueoccurs as fibrous bundles, with properties varying widely from thenormal host tissue, and the structure of the implant device in thisembodiment will tailor the growth of the scar tissue such that itsproperties will approach that of the native tissue. The structure of theimplant is used to train the tissue, such that scar tissue forms in anon-bundled form (e.g., fibrous strands, more linear arrays, or smalleror thinner bundles), and the structure has enough integrity to supportthe growing tissue such that it does not contract non-uniformly, therebyavoiding or minimizing the disfiguring characteristics caused byshrinking of the tissues during final stages of growth and/or bundling.Additionally, this physical or geometric modeling of tissue may be aidedby the delivery of a targeted therapy.

In the foregoing embodiments, it is envisioned that therapy delivery maybe by way of incorporation of the therapy into the device matrix,macrostructure, microstructure, or microspheres (regardless of wherelocated), and regardless of whether the therapy was delivered uniformly,time-staged, or as a burst dose. These methods of therapy delivery arelocalized in nature, as opposed to systemic approaches, that arenecessarily delivered via the blood-stream. These systemic approachesconcomitantly deliver therapies to various tissue and organs for whichthey were not intended. Localized delivery may allow higher doses, atthe target site, than are tolerable to the body as deliveredsystemically. Chemotherapeutic treatment for certain cancers as well asother diseases may particularly be amenable to this type of therapydelivery, although various other procedures, not limited to those inTable 1, may benefit. Secondary therapies, or therapies deliveredsimultaneously with primary therapies, may be beneficial to reduce oreliminate side-effects of the primary therapy.

It is envisioned that time-staged delivery, whether achieved by apreferred placement of therapy within the macrostructure,microstructure, or microsphere, would allow staging of treatment, one ofwhich stages may actually be detrimental to cell growth andproliferation, prior to the delivery of therapies that aid in tissueingrowth or regeneration. Furthermore, tissue ingrowth and regenerationmay have stages, such as, the initial nurturing therapy followed byrapid growth and proliferation aids.

As an example, Cisplatin and Paclitaxel are commonly used together inchemotherapeutic applications. These embodiments could deliverPaclitaxel at high dose rates initially, followed by lower dose rates ofCisplatin, which would occur over longer periods of time. It is alsoenvisioned by this invention that the first therapy may be housed in amicrostructural element (e.g., Paclitaxel) while the second therapy(e.g., Cisplatin) is housed in the matrix macrostructure. The slowerresorbing macrostructure would supply the localized dose of the secondtherapy over the entire time during which any of the macrostructureremained.

In yet another embodiment, time-staged delivery or secondary therapydelivery may allow the function of tissue (e.g., organ such as theliver, etc.) to be replaced or supported, prior to, or concurrent with,regrowth or regeneration of diseased or removed tissue, or cellulartransplant, which may be accomplished by the foregoing embodiments. Thissupport may allow the tissue to slowly regain organic function, orreassume total function, whereas the otherwise diminished capacity maylead to total organ failure. Additionally, this support function therapymay be utilized to counteract a side effect of the primary therapy. As anon-limiting example, it may be used to support liver function duringchemotherapy. The aforementioned localized delivery, together withsecondary support, may allow the use of drugs not otherwise tolerated,or current drugs in greater dosages.

This type of cellular transplant embodiment may incorporate cells in anyof the various regions, as disclosed in the other embodiments, or othersites within the implant (e.g., macrostructure, microstructure, voidspace, or microsphere). Additionally, therapies may be located in any ofthese regions.

TABLE 4 Examples with Some Sub-types of Biological, Pharmaceutical, andother Therapies Deliverable via the Device in Accordance with thePresent Invention Adenovirus with or without genetic material Angiogenicagents Angiotensin Converting Enzyme Inhibitors (ACE inhibitors)Angiotensin II antagonists Anti-angiogenic agents AntiarrhythmicsAnti-bacterial agents Antibiotics Erythromycin PenicillinAnti-coagulants Heparin Anti-growth factors Anti-inflammatory agentsDexamethasone Aspirin Hydrocortisone Antioxidants Anti-platelet agentsForskolin Anti-proliferation agents Anti-rejection agents RapamycinAnti-restenosis agents Antisense Anti-thrombogenic agents ArgatrobanHirudin GP IIb/IIIa inhibitors Anti-virus drugs Arteriogenesis agentsacidic fibroblast growth factor (aFGF) angiogenin angiotropin basicfibroblast growth factor (bFGF) Bone morphogenic proteins (BMP)epidermal growth factor (EGF) fibrin granulocyte-macrophage colonystimulating factor (GM-CSF) hepatocyte growth factor (HGF) HIF-1 Indianhedgehog (Inh) insulin growth factor-1 (IGF-1) interleukin-8 (IL-8)MAC-1 nicotinamide platelet-derived endothelial cell growth factor(PD-ECGF) platelet-derived growth factor (PDGF) transforming growthfactors alpha & beta (TGF-.alpha., TGF- beta.) tumor necrosis factoralpha (TNF-.alpha.) vascular endothelial growth factor (VEGF) vascularpermeability factor (VPF) Bacteria Beta blocker Blood clotting factorBone morphogenic proteins (BMP) Calcium channel blockers CarcinogensCells Stem cells Bone Marrow Blood cells Fat Cells Muscle CellsUmbilical cord cells Chemotherapeutic agents Ceramide Taxol CisplatinPaclitaxel Cholesterol reducers Chondroitin Clopidegrel (e.g., plavix)Collagen Inhibitors Colony stimulating factors Coumadin Cytokinesprostaglandins Dentin Etretinate Genetic material GlucosamineGlycosaminoglycans GP IIb/IIIa inhibitors L-703,081Granulocyte-macrophage colony stimulating factor (GM-CSF) Growth factorantagonists or inhibitors Growth factors Autologous Growth FactorsBovine derived cytokines Cartilage Derived Growth Factor (CDGF)Endothelial Cell Growth Factor (ECGF) Epidermal growth factor (EGF)Fibroblast Growth Factors (FGF) Hepatocyte growth factor (HGF)Insulin-like Growth Factors (e.g. IGF-I) Nerve growth factor (NGF)Platelet Derived Growth Factor (PDGF) Recombinant NGF (rhNGF) Tissuenecrosis factor (TNF) Tissue derived cytokines Transforming growthfactors alpha (TGF-alpha) Transforming growth factors beta (TGF-beta)Vascular Endothelial Growth Factor (VEGF) Vascular permeability factor(UPF) Acidic fibroblast growth factor (aFGF) Basic fibroblast growthfactor (bFGF) Epidermal growth factor (EGF) Hepatocyte growth factor(HGF) Insulin growth factor-1 (IGF-1) Platelet-derived endothelial cellgrowth factor (PD-ECGF) Tumor necrosis factor alpha (TNF-.alpha.) Growthhormones Heparin sulfate proteoglycan HMC-CoA reductase inhibitors(statins) Hormones Erythropoietin Immoxidal Immunosuppressant agentsInflammatory mediator Insulin Interleukins Interlukins Interlukin-8(IL-8) Lipid lowering agents Lipo-proteins Low-molecular weight heparinLymphocites Lysine MAC-1 Morphogens Bone morphogenic proteins (BMPs)Nitric oxide (NO) Nucleotides Peptides PR39 Proteins ProstaglandinsProteoglycans Perlecan Radioactive materials Iodine-125 Iodine-131Iridium-192 Palladium 103 Radio-pharmaceuticals Secondary MessengersCeramide Signal Transduction Factors Signaling Proteins SomatomedinsStatins Stem Cells Steroids Thrombin Sulfonyl Thrombin inhibitorThrombolytics Ticlid Tyrosine kinase Inhibitors ST638 AG-17 VasodilatorHistamine Forskolin Nitroglycerin Vitamins E C Yeast

Also within the inventive concept of the present invention is theplacing of a plurality of microstructure materials at strategiclocations within the same implant to perform multiple and variedbiologic functions. For example, a large osteochondral defect maybenefit from hyaluronan velour for microstructure in the subchondralregion intended for osteoneogenesis. The placement of a differentmicrostructure material can be accomplished by various methods,including investing the microstructure material into the regions beforethey are joined, by investing the device or regions thereof beforejoining from a first surface with a desired volume of microstructurematerial less than the total void volume of the macrostructure and theninvesting from the opposite surface with a volume of a differentmicrostructure material equal to the balance of void volume of themacrostructure.

Except for the critical location at the interface between the cartilageregion (or first region, where applicable), the material of thesubchondral bone region (or second region, where applicable) ishydrophilic by virtue of being treated with a wetting agent such as setforth in U.S. Pat. No. 4,186,448. For example, beginning at about 200 to1500 micrometers, but more preferably 500 to 800 micrometers, from theinterface surface and extending into the subchondral bone region, themacrostructure polymer of the subchondral bone region may be renderedhydrophobic, such as by treating the entire device or the subchondralbone region with a surfactant and then inactivating the surfactant inthe hydrophobic barrier region (i.e., between its interface with thefirst and second regions or macrostructures), or by not treating thebarrier surfaces with a surfactant while the remaining portions aretreated.

Likewise, a hydrophobic barrier may be created within a device of simple(i.e. single) or complex (i.e. multiple) internal architectures by othermeans. For example, a separate fibrillar construct of bioresorbablepolymer may be fabricated devoid of surfactant and may be interspersedbetween two segments of a device whose polymers have been renderedhydrophilic.

For example, in a simple device, such as one used to create cartilageand bone, the bone regeneration region (e.g., alpha-hydroxy-acid) isabout 40 to about 90 percent of the apparent volume of the device, withthe barrier located between the bone and cartilage regions. It isrecognized that the barrier, as described above, may be a materialdistinct from the first and second regions, or it may exist at or nearthe surface of one of the regions, prior to the joining of the regions.

Furthermore, the barrier may, in a preferred embodiment, compriseinterdigitations of the two joined regions.

In certain applications, it is envisioned that a total fluid or liquidbarrier is a necessity, while other applications may have some toleranceor even a need for some liquid through-flow. The type and amount(quantity per application or number of applications) of surfactant cangreatly influence the effectiveness of the barrier's inhibition ofliquid flow interference. This invention contemplates a barrier thatallows no fluid flow, as well as some small amount or retarded flowrate. This entire range of flow being referred to as “inhibited.”

Additionally, the term surfactant, as used herein, envisions traditionalionic and stearic treatments, as well as dissimilar material coatings,utilized to alter the host material's response to water and/or certainother liquids. For example, it is envisioned that a hydrophilic coatingmay be applied to a hydrophobic structure or substrate, therebyrendering the body, or section thereof, hydrophilic, and vice versa.

Alternatively, other surface or chemical modification techniques may beutilized to create a suitable barrier between adjacent regions, or as anintraregional barrier. Such techniques include but are not limited toion-beam activation, plasma, radio frequency, ultrasound, radiation, andthermal processing.

Water-based fluids, specifically fluid blood, brought to this locale bycapillary action through hydrophilic polymer of the subchondral boneregion closest to subehondral bone, are prohibited from travelingfurther toward the cartilage region by a hydrophobic polymer of thesubehondral bone region in this vicinity. The interstices of thehydrophobic fibrillar membrane would eventually accommodate cell growthinto, and/or migration through, the hydrophobic zone, but the immediateeffect of such a membrane would be to prevent passage of water-basedfluids across its boundaries.

The hydrophobic barrier is a significant advance and development fordevices intended for use in chondroneogenesis, because hyalinecartilage, specifically the articular cartilage of joints, is anavascular tissue and must be protected from intimate contact withsources of high oxygen tension such as blood. When the recipientcartilage tissue defect is prepared to receive the implant, it isnecessary to continue the defect into the underlying subchondral bone,called the cancellous bone, to assure that there will be new bone formedbeneath the cartilage region which will produce a competent bond withthe newly developing cartilage.

The customization of a microenvironment has been disclosed, wherein athree-dimensional architecture may support cell growth. However, theapproach was that of a modified cellular structure, not a physical orgeometric attribute (Grande, et. al.: A dual gene therapy approach toosteochondral defect repair using a bilayer implant containing BMP-7 andIGF-1 transduced periosteal cells. 47.sup.th Annual Meeting, OrthopaedicResearch Society, Feb. 25-28, 2001, San Francisco, Calif.). Thistechnique differs from the present invention as it does not include anysubchondral bone sector. Therefore, complete natural bonding betweenbone and cartilage of sufficient integrity remains problematic.

A similar technique has recently been disclosed, which includes apartition of the microenvironments (Gao J. et. al.: Tissue engineeredosteochondral graft using rat marrow-derived mesenchymal stem cells.47.sup.th Annual Meeting, Orthopaedic Research Society, Feb. 25-28,2001, San Francisco, Calif.). That construct has two regions gluedtogether with fibrin glue. The regions comprise insoluble hyaluronicacid and tricalcium phosphate. The drawback of this construct is thatthe barrier is hydrophilic. Additionally, the fibrin glue is quicklybioresorbable and lacks significant adhesive strength. Further,hydrophilic barriers of this construct allow the transport of bodyfluids and soluble cytokines between regions, which interruptschondrogenesis and osteogenesis. Barriers which are quicklybioresorbable promote unstable interfaces resulting in mechanical andbiological insufficiencies.

Tissue preparation, such as this, engages the rich collateralcirculation of subchondral cancellous bone and its associated bonemarrow. If the cultured chondrocytes or host cartilage cells come intocontact with the fluid blood produced by this source of collateralcirculation, they will fail to maintain their chondrocyte phenotype.However, the hydrophobic barrier as may be employed in the presentinvention described above isolates the cartilage region from contactwith whole blood originating in the subchondral bone region. Thistissue-specific construct is exemplary, as other regions or tissues andother fluids are contemplated.

It can be appreciated that an anatomically specific device, which may bebioresorbable, according to the teachings of the foregoing inventionshaving a fabricated macrostructure closely resembling the mature tissueswhich are to be regenerated by the completed implant, has particularvalue. Further, integrating one or more of a macrostructure,microstructure, cells cultured in vitro, culture medium and associatedgrowth factors, morphogens, drugs and other therapeutic agents mayadditionally be beneficial.

According to the teachings of the present invention, the device can beutilized as a transport system for chondrocytes, growth factors,morphogens and other biologically active agents, in treatment ofarticular cartilage defects. Suitable source tissue is harvested, andthe cells are cultured using standard chondrocyte culturing methods,with the specific cell type in the preferred form being articularcartilage chondrocyte. The cartilage defect is surgically prepared byremoving diseased or damaged cartilage to create a cartilage andsubchondral bone defect, with the defect extending approximately 0.5 cmto 1.0 cm into subchondral cancellous bone. With the device and defecthaving generally the same shape, the device is inserted into the tissuedefect such as by press fitting. A volume of in vitro cell culturesuspension is measured out by a microliter syringe which generallymatches exactly the void volume of the cartilage region macrostructureinvested by the microstructure and is injected onto the outer surface ofthe tangential zone of the cartilage region and which will ultimately bein contact with synovial fluid. The joint anatomy can then be replacedin proper position and the wound can be closed.

Alternatively, cells or other therapeutic additives may be incorporatedduring the manufacture of the device, or during the final devicepreparation (i.e., immediately prior to implant), or as briefly notedabove following the implant procedure (e.g., prior to wound closure oras a later therapy, following wound closure).

Although the preferred form relates to the transport and/or in vivoculturing of chondrocytes, it should be noted that the teachings of thepresent invention, and the useful devices fabricated as a resultthereof, are intended to culture and/or transport, and to sustain inlife, any cell type having therapeutic value to animals and plants.Various other cell types would be beneficial for tissue other thancartilage or bone, depending on the site and application. The varioususes outlined above in text and tabular form are contemplated by thisinvention.

The term “therapy” has been used in this specification, in variousinstances. Notwithstanding these various uses, many in combination withother agents (e.g., drug, biologic, biologically active agents, etc.),therapy is not meant to be exclusive of these, but rather to incorporatethem. The usage herein is employed to be more descriptive of potentialtreatment forms, and not limiting as to the definition of the term.

Thus since the invention disclosed herein may be embodied in otherspecific forms without departing from the spirit or generalcharacteristics thereof, some of which forms have been indicated, theembodiments described herein are to be considered in all respectsillustrative and not restrictive. The scope of the invention is to beindicated by the appended claims, rather than by the foregoingdescription, and all changes which come within the meaning and range ofequivalency of the claims are intended to be embraced therein.

1. An implant device that provides a biologically acceptable andmechanically stable surface structure for regeneration of tissue, saidimplant device comprising; a. a first region comprising a regenerationcomplex comprising at least one resorbable polymer; b. a second regioncomprising a regeneration complex comprising collagen; and c. a zonecomprising a hydrophobic barrier.
 2. The implant device of claim 1,wherein the second region is arranged to accept the ingress of cells,and having mechanical or physical properties which affect cellularattachment, thereby affecting cell phenotype.
 3. The implant device ofclaim 2, wherein said affecting cellular attachment is caused byinteraction between said second region and endocellular fibrils of saidcells.
 4. The implant device of claim 1, wherein said hydrophobicbarrier is arranged at an interface between said first and secondregions.
 5. The implant device of claim 4 wherein said hydrophobicbarrier is arranged as interdigitations between the first and secondregions.
 6. The implant device of claim 1, wherein said first region isarranged as a subchondral bone region, and the second region is arrangedas a cartilage region.
 7. The implant device of claim 1, wherein saidresorbable polymer regeneration complex comprises a firstmacrostructure, and said collagen regeneration complex comprises aporous fabric.
 8. The implant device of claim 1, wherein at least one ofsaid first region and said second region comprises complex internalarchitectures.
 9. The implant device of claim 4, wherein saidhydrophobic barrier comprises a fibrillar membrane arranged to preventthe passage of water-based fluids across the membrane.
 10. The implantdevice of claim 9, wherein said membrane acts as a barrier to thepassage of water based fluids at least temporarily.
 11. The implantdevice of claim 9, wherein said hydrophobic membrane is arranged toallow the ingrowth of cells into and through said hydrophobic membrane.12. The implant device of claim 9, wherein said hydrophobic barrier isarranged to 45 prevent the passage of blood therethrough therebycreating an avascularized region.
 13. The implant device of claim 9,wherein said hydrophobic barrier is arranged to prevent the passage ofblood across the membrane thereby creating, on one side of the barrier,an avascularized region, and, on the other side of the barrier, avascularized region.
 14. The implant device of claim 7, wherein saidporous fabric comprises a second macrostructure comprising abiologically acceptable polymer arranged as a unitary porous body havingenclosed pores that are random in size, position and shape, such poresforming interconnecting voids in communication with the exterior of saidporous body.
 15. The implant device of 14, with said porous fabrichaving physical properties arranged to influence interactions betweensaid second macrostructure and endocellular fibrils associated with saidsecond macrostructure.
 16. The implant device of claim 14, wherein saidinterconnecting voids of said second macrostructure are arranged toinfluence forms of regenerated tissue.
 17. The implant device of claim14 herein said second macrostructure is arranged to mimic naturalchondrocyte host environment and facilitate regenerative cascades tooccur for replication of damaged tissue.
 18. The implant device of claim14, wherein said second macrostructure further comprise a microstructurearranged within the interconnecting voids of the pores of the secondmacrostructure.
 19. The implant device of claim 18, wherein saidmicrostructure is arranged to affect the degree of matrix matching. 20.The implant device of claim 1, wherein said regeneration complex furthercomprises at least one additive selected from the group consisting of:composite elements, biologics, pharmaceuticals, or therapies.
 21. Theimplant device of claim 20 wherein said additive is a composite elementcomprising ceramic.
 22. The implant device of claim 21 wherein theceramic is selected from at least one of the group consisting of calciumphosphate, calcium sulfate and tricalcium phosphate.
 23. The implantdevice of claim 20 wherein the additive is selected from at least one ofthe group consisting of antiobiotics, anti-coagulants, anti-rejectionagents, arteriogenesis agents, cells, chemotherapeutic agents, growthfactors, hormones, morphogens or vitamins.
 24. The implant device ofclaim 20, wherein said additive material is added to said secondmacrostructure.
 25. A porous, multi-component device for repair ofarticular cartilage defects, comprising: a. a first region comprising afirst component in the form of a microstructure presenting a porousintercommunicating network for cellular attachment, and a secondcomponent in the form of a fibrous macrostructure supporting the firstcomponent and; b. a second region comprising a third component of porouspolymer attached to the first and second components.
 26. The device ofclaim 25, wherein said first component comprises acid soluble collagen.27. The device of claim 25, wherein said second component comprisesinsoluble collagen.
 28. The device of claim 25, wherein said polymer ofsaid third component comprises polymerized alpha-hydroxy acids.
 29. Thedevice of claim 25, wherein said third component further comprises atleast one ceramic.
 30. The device of claim 25, further comprising afourth component arranged at an interface between said first and secondregions.
 31. The device of claim 30, wherein said fourth componentcomprises a film.
 32. The device of claim 31, wherein said film isporous.
 33. The device of claim 31, wherein said film is hydrophobic.34. The device of claim 30, wherein said first region comprises asecurement complex for the device, said second region comprises aregeneration complex, and said fourth component comprisesinterdigitations between the first and second regions.
 35. The device ofclaim 25, wherein said device further comprises at least one additiveselected from the group consisting of: composite elements, biologics,pharmaceuticals, or therapies.
 36. The device of claim 35 wherein saidadditive is a composite element comprising ceramic.
 37. The device ofclaim 36 wherein the ceramic is selected from at least one of the groupconsisting of calcium phosphate, calcium sulfate and tricalciumphosphate.
 38. The device of claim 35 wherein the additive is selectedfrom at least one of the group consisting of antiobiotics,anti-coagulants, anti-rejection agents, arteriogenesis agents, cells,chemotherapeutic agents, growth factors, hormones, morphogens orvitamins.
 39. An implantable regeneration complex for the healing oftissue, where such complex provides a network of voids wherein fluidsabsorbed into the voids allows the complex to take on properties thatmimic host tissue in the region of the complex.
 40. The implantableregeneration complex of claim 39, wherein said fluids are absorbed froma source comprising the adjacent host tissue.
 41. The implantableregeneration complex of claim 39, wherein said properties are selectedfrom at least one of the group consisting of mechanical, chemical orbiological.
 42. The implantable regeneration complex of claim 39,wherein said absorbed fluids further provide securement of said complexin said host tissue.
 43. A porous regeneration complex for the healingof tissue, where such complex provides a network of voids wherein fluidsabsorbed into the voids alters the physical properties of the complex,such that the complex has properties similar to the host tissue.
 44. Theporous regeneration complex of claim 43, wherein said properties areselected from at least one of the group consisting of mechanical,chemical or biological.
 45. A porous three-component regenerationcomplex for repair of articular cartilage defects comprising: a. a firstcomponent of rapidly degrading polymer that swells as it imbibeswater-based fluids; b. a second component of polymer associated with thefirst component that entraps the water-based fluids and said swollenfirst component; and c. a third component of polymer attached to saidfirst and second components, said third component being in the form of amatrix matched complex; wherein the fluids absorbed into the complexallows the complex to take on properties that mimic host tissue in theregion of the implanted complex.
 46. The regeneration complex of claim45 wherein the first component is composed of acid soluble collagen. 47.The regeneration complex of claim 45 wherein the second component iscomposed of insoluble collagen fibers.
 48. The regeneration complex ofclaim 45 wherein the third component is composed of a polymerizedalpha-hydroxy acid.
 49. The regeneration complex of claim 45 wherein thethird component additionally contains a calcium mineral.